The medium contains 1.0% each of
sucrose and lactose and 0.1% glucose. If only
glucose is fermented, acid produced in the butt will turn it yellow,
but insufficient acid products are formed to affect the methyl red in
the slant. However, if either sucrose or lactose are fermented,
sufficient fermentation products will be formed to turn both the butt and
the
slant yellow. If gas is formed during the fermentation, it will show in
the butt either as bubbles or as cracking of the agar. If no
fermentation occurs (as for an obligate aerobe), the slant and
butt will remain red.The medium
also contains ferrous sulfate. If the bacterium forms H2S,
this chemical will react with the iron to form ferrous sulfide, which
is seen as a black precipitate in the butt (a black butt). The
following table summarizes these reactions:

Phenol red turns yellow in an
acid environment. It thus indicates
whether the acids of fermentation have been produced. Failure to
turn the butt yellow
indicates that no fermentation has occured, and that the bacterium is
an obligate aerobe.

0.1 % glucose

if only glucose is fermented,
only a small amount of acid is produced

If only glucose is fermented,
only enough acid is produced to turn the butt yellow. The slant
will remain red.

1.0 % lactose
1.0% sucrose

if the culture can ferment
either lactose (lac+) and/or sucrose (suc+), a large amount of
acid is produced

a large amount of acid turns
both butt and slant yellow, thus
indicating the ability of the culture to ferment either lactose or
sucrose

FeSO4
(ferrous sulfate)

A source of iron and sulfur

A few bacteria are capable of
reducing the SO4= to H2S (hydrogen sulfide).
The iron combines with the H2S to form FeS (ferrous sulfide) a black
compound. This will turn the butt black. Thus, a black butt
indicates
H2S production.

Here are six different reactions for bacteria inoculated into
triple sugar iron slants.
Look at the left image first and record what you think the reactions
are in each slant. Then check your answers with the image on the
right.

After you have scored your TSI Agar
slants, you should suggest a species of bacterium which matches those
metabolic traits discovered. While by no means definitive, the
following are TSI Agar reactions typical of a number of prominent
species which can be distinguished with this medium: